In the mammalian central nervous system, L-glutamate serves as a major excitatory neurotransmitter. The interaction of glutamate with its membrane-bound receptors is believed to play a role in many important neuronal processes, including, for example, fast synaptic transmission, synaptic plasticity and long-term potentiation. These processes are fundamental to the maintenance of life and normal human abilities such as learning and memory. Monaghan D. T. et al., 8 Neuron 267 (1992).
Pharmacological characterization of receptors for L-glutamate has led to their classification into two families based on their biological function: the ionotropic receptors which are directly coupled to cation channels in the cell membrane, and the metabotropic receptors which function through coupling to G-proteins. A number of ionotropic receptors have been further characterized on the basis of the relatively specific agonists by which they can be activated. One major group comprises those receptors activated by N-methyl-D-aspartate (NMDA), which appears to have multiple allosteric modulatory sites. The other two groups consist of those receptors activated by kainate and/or amino-3-hydroxy-5-methyl-4-isoxozole propionate (AMPA). Collingridge G. L. et al., 40 Pharmacol. Rev. 143 (1989).
Molecular cloning studies of rodent ionotropic receptors have recently provided some information on the molecular structure of these proteins. The cDNAs for seven different subtypes of the kainate/AMPA group have been characterized. Heinemann S. et al., PCT publication, W091/06648 (1991), Keinanen K. et al., 249 Science 556 (1990), Sakimura K. et al., 272 FEBS Lett. 73 (1990), Werner P. et al., 341 Nature 742 (1991), Bettler B. et al., 8 Neuron 257 (1992). Splice variants, referred to as "flip" and "flop", of some of these have been characterized as well. Sommer B. et al., 249 Science 1580 (1990). In addition, one member of the NMDA group has been cloned. Moriyoshi, K. et al., 354 Nature 31 (1991). An NNDA-related protein has also been reported. Kumar K. N. et al., 354 Nature 70 (1991). These proteins share varying degrees of homology with one another and are therefore believed to represent a gene superfamily. Based on analogy with other better characterized ion channel receptors, glutamate ionotropic receptors are expected to exist in vivo within the cell membrane as multisubunit assemblies of these subunits. Unwin N., 3 Neuron 665 (1989).
Moreover, at least two human glutamate receptors have been reported as cloned. The reported human receptors differ slightly from the present invention. Puckett C. et al., 88 Proc. Nat. Acad. Sci. 7557 (1991) and Sun W. et al., 89 Proc. Nat. Acad. Sci. 1443 (1992). The glutamate receptor cloned by Puckett et al. was named GluHI and was later identified to be the "flip" version of this particular receptor. The Sun W. et al. reference refers to the glutamate receptor they cloned as the HBGR1 receptor and explains that HBGR1 is presumed the "flop" version of GluHI. Sun et al. also discloses a partial clone of HBGR2, or GluH2.
In addition to its role in normal human physiology, interaction of L-glutamate with its receptors is believed to play a key role in many neurological disorders such as stroke, epilepsy and head trauma, as well as neurodegenerative processes such as Alzheimer's disease. Olney R. W., 17 Drug Dev. Resa., 299 (1999). For this reason, understanding the molecular structure of human L-glutamate receptors will be important for understanding these disease processes as well as furthering the search for effective therapeutic agents. Up to the present, the search for therapeutic agents which will selectively bind and modulate the function of human glutamate receptors has been hampered by the unavailability of homogeneous sources of receptors to use for screens and tests of potential drug candidate compounds. The brain tissues commonly used by pharmacologists presently are derived from experimental animals (non-human) and furthermore contain mixtures of various types of glutamate receptors.
In searching for drugs for human therapy it is desirable to use receptors that are more analogous to those in the intact human brain than are the rodent receptors employed to date. The current invention provides a human receptor which can be used to search for drugs which modulate these receptors.